Figure 2
Characterization of proviral clones in vitro. 293 T cells (2 × 105) were co-transfected with 1 μg of wtHTLV-2 or HTLV-2Δp28 proviral clones or negative control DNA along with 0.1 μg of LTR-1-Luc and 0.01 μg of TK-Renilla. All transfections were performed in triplicate and normalized to TK-Renilla to control for transfection efficiency. Cell lysates or supernatants were harvested 48 h post-transfection. (A) Measure of Tax activity presented as relative luciferase units. Results indicated that loss of p28 expression from the proviral clone did not significantly alter Tax activity. (B) Rex activity as measured by expression of p19 Gag (virions) in the cellular supernatants. Results indicated that loss of p28 expression from the proviral clone did not significantly alter Rex activity. (C) Total RNA was extracted from 293 T cells transfected with HTLV-2 or HTLV-2pΔ28 as in panels A and B. mRNA copy number was quantified by Taqman realtime RT-PCR. The histogram represents the copy number of gag-pol, tax/rex, and p28 transcripts normalized to 1 × 106 copies of gapdh. Results indicated that deletion of p28 protein had no significant affect on tax/rex, gag/pol, or p28 mRNA expression.